I. Technical Field
The present invention relates to a cell culturing method employing biomechanical stimulation loading and its system for applying, in in vitro culture, repeated biomechanical stimulation to culture cells, culture tissues and a construct including the culture cells (which are collectively referred to as “cultures” in the description). The repeated biomechanical stimulation is stimulation similar to vertical weight load stimulation or sideward shear stress stimulation which acts on tissues in an aboveground living body.
II. Description of the Related Art
As an approach to treating patients with cartilage lesions, cell transplantation therapy has been recently carried out, for example, in Europe and America. In this approach, remaining healthy cartilage tissues are taken out from the patients, cartilage cells are separated from the tissues and undergo in vitro culture for growth, and cultured cartilage cells are transplanted back to the patients.
This approach, which carries out the in vitro cartilage cell culture and growth, however has problems that it does not carry out in vitro regeneration of cartilage tissues, produces an unstable effect and requires long-term repair and maturation of the cartilage tissues.
In the living body, tissues and cells are subjected to not only biochemical stimulation but also mechanical stimulation attributed to gravity, muscle contraction force or external force. Culture cells and culture tissues are known to change their functions including growth, differentiation and metabolism by experiencing biomechanical stimulation in static or dynamic form.
Based on such findings, cell and tissue culture systems utilizing the biomechanical stimulation have been recently studied and put to practical use by several domestic and foreign research groups, including, for example, a system disclosed in Japanese Patent Unexamined Publication No. 2003-265164.
Culture systems using stress stimulation include a method of expanding and contracting or shaking a membrane member to which the cells are adhered for culturing the cartilage cells or vascular smooth muscle cells, and a method of placing a support holding the cartilage cells in a column and applying the stress stimulation to the cartilage cells held by the support by means of the flow of culture medium or culture fluid circulated by a pump for culturing the cartilage cells.
Culture systems using hydrostatic pressure stimulation include a method of applying pressure to the cartilage cells by means of a gas cylinder for stimulation loading and a method of pressurizing the column with a 5 MPa load, which is equivalent to intra-articular pressure, by means of a hydraulic cylinder pump for stimulating the cartilage cells.
There has also been proposed a cell culture system in which the culture medium is fed into the column (that is, to a device in the column) by a pump and the flow rate of the culture medium is varied by operation of a valve or the like for desired pressure loading patterns and cycles.
These methods and systems re-create conditions close to those in the living body, such as levels of pressure on the cells in the living body, pressure variations and patterns in pressure rise and drop, through use of the culture fluid. However, these methods and systems are strictly for the purpose of stimulating the culture cells by expanding the culture cells and using the flow of culture fluid and the culture fluid, not re-creating vertical biomechanical stimulation, which is attributed to gravity, to support tissues of a bone, a cartilage or the like.
As is obvious from the known fact that healthy astronauts who experience reduced gravity stimulation in outer space of microgravity and patients or aged persons who have to stay on beds develop osteoporosis at their spines, arms and legs, the vertical repeated stimulation attributed to gravity is of importance to the living body.